Temperature-sensitive vaccinia virus mutants identify a gene with an essential role in viral replication.
Vaccinia virus mutants ts2 and ts25, members of the same complementation group, exhibit a temperature-dependent arrest at the stage of viral DNA replication. The lesions responsible for the mutant phenotypes have been localized to the far left region of the HindIII B genomic fragment by marker rescue studies. Hybrid selection analyses established that the DNA fragments positive for rescue represented the first open reading frame of the HindIII B fragment and encoded a 30-kilodalton protein. The gene is expressed early after infection as a rightwardly transcribed 1-kilobase-pair mRNA whose coordinates were determined by S1 nuclease mapping. To further the phenotypic analysis of the mutants, the accumulation of viral DNA sequences during permissive and nonpermissive infections was quantitated. The extent of the DNA- phenotype was shown to vary in different cell types. In mouse L cells at either high or low multiplicity of infection, nonpermissive DNA synthesis was less than 5% of that seen in permissive infections. This severe defect was mirrored by correspondingly low viral yields. In infections of BSC40 monkey cells, however, the deficiencies in both DNA synthesis and virus production were far less severe. For one mutant (ts2), the temperature sensitivity in BSC40 cells varied inversely with the multiplicity of infection.
Rempel
R E
RE
Department of Cell Biology, Cornell University Medical College, New York, New York 10021.
Anderson
M K
MK
Evans
E
E
Traktman
P
P
D000818:Animals; D002460:Cell Line; D003360:Cosmids; D004279:DNA, Viral / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D005814:Genes, Viral; D007700:Kinetics; D007739:L Cells; D051379:Mice; D008970:Molecular Weight; D009154:Mutation; D009693:Nucleic Acid Hybridization; D010957:Plasmids; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000235:genetics; D012367:RNA, Viral / Q000235:genetics; D015183:Restriction Mapping; D013696:Temperature; D014616:Vaccinia virus / Q000235:genetics* / Q000502:physiology; D014764:Viral Proteins / Q000235:genetics; D014779:Virus Replication
D004279:DNA, Viral; D012333:RNA, Messenger; D012367:RNA, Viral; D014764:Viral Proteins